Nystatin—dextran conjugates: Synthesis and characterization

The coupling of nystatin (Nys), a water-insoluble antifungal agent, to dextran via an imine or amine bond was systematically investigated. Dextran was first oxidized to dialdehyde dextran using potassium periodate, purified from the oxidizing agent, and reacted with Nys to form the Schiff base. The Schiff base was reduced to the amine using borohydride. All reactions took place in water. The purification of the oxidized dextran from the oxidizing agent was essential to prevent oxidative degradation of Nys at the coupling step. The effects on the coupling yield of the following factors: dextran molecular weight, degree of oxidation (aldehyde content), Nys to dextran ratio, temperature, and reaction pH were studied. A 95% coupling yield was obtained at the optimized coupling conditions: pH 8.9 ± 0.1, 50% degree of oxidation, and initial ratio of Nys to dialdehyde dextran 1:2.5. In all experiments, dextran was decreased in molecular weight during the oxidation step. Both imine and amine forms of Nys-dextran conjugates were soluble in water and exhibited improved stability in aqueous solutions as compared to the unbound drug. The conjugates showed comparable minimum inhibitory concentration (MIC) values against Candida albicans and Cryptococcus neoformans. The conjugates were about 25 times less toxic than free Nys after a single injection in mice. © 1996 John Wiley & Sons, Inc.     

Polyamidoamine dendrimer and dextran conjugates: preparation,characterization, and in vitro and in vivo evaluation

Amide and ester conjugates of aceclofenac with polyamidoamine (PAMAM-G0) dendrimer zero generation and dextran (40 kDa) polymeric carrier, respectively, are presented. The prepared conjugates were characterized by UV, TLC, HPLC, IR, and ¹H NMR spectroscopy. The average degrees of substitution of amide and ester conjugates were determined and found to be (12.5 ± 0.24) % and (7.5 ± 0.25) %, respectively. The in vitro hydrolysis studies showed that dextran ester conjugate hydrolyzed faster in a phosphate buffer solution of pH 9.0 as compared to PAMAM dendrimer G0 amide conjugate, and followed the first order kinetics. No amount of the drug was regenerated at pH 1.2 in simulated gastric fluid. The dextran conjugate showed short half-life as compared to the PAMAM dendrimer conjugate. Anti-inflammatory and analgesic activities of the dendrimer conjugate were found to be similar to those of the standard drug. Results of chronic ulceroginic activity showed deep ulceration and high ulcer index for aceclofenac, whereas lower ulcer index was found for the PAMAM dendrimer and dextran (40 kDa) conjugates. Experimental data suggest that PAMAM dendrimer and dextran (40 kDa) can be used as carriers for the sustained delivery of aceclofenac along with a remarkable reduction in gastrointestinal toxicity.     

Molecular characteristics and antioxidant activity of polyethylene glycols modified by sterically hindered phenols

Chemical modification of polyethylene glycols by antioxidants belonging to the class of sterically hindered phenols is used to obtain water-soluble conjugates differing in the structure of the joined antioxidant and molecular weight. The inclusion of hydrophobic end groups in polyethylene glycol molecules leads to a decrease in the lower critical mixing point of the solution as compared to the original polymer. Dilute solution viscometry and light scattering are used to determine the molecular-mass characteristics of the polymers and the hydrodynamic radii of single conjugate molecules. The mass fraction of single molecules in aqueous solutions of the conjugates is greater than 95%. The aggregates are micellar-type particles whose core is formed by the hydrophobic moieties of sterically hindered phenols. It is shown that the antiradical activity of the antioxidants 3-(3-tert-butyl-4-hydroxy-5-methylphenyl)-propionic acid and 3- (3,5-di-tert-butyl-4-hydroxyphenyl)-propionic acid, which are used for the modification, differs only slightly, whereas the activity of the relevant conjugates increases significantly.     

Fe3O4/葡聚糖/抗体磁性纳米生物探针的制备和层析检测

在免疫检测中 ,经常利用一些具有特殊物理化学性质的标记物对抗体 (或抗原 )进行偶联标记 ,在抗体与抗原识别后 ,通过对标记物的定性和定量检测而达到对抗原 (或抗体 )检测的目的 .传统的免疫标记物包括放射性同位素 [1] 、酶 [2 ] 、胶体金 [3] 和有机荧光染料分子 [4 ] 等 .近年来 ,随着纳米技术的发展 ,半导体荧光纳米晶 [5,6 ] 和磁性纳米晶 [7] 在免疫检测方面受到了广泛关注 .磁性纳米晶性能稳定 ,较易制备 ,可与多种分子复合使粒子表面功能化 ,并且由于磁纳米晶具有超顺磁性 ,为样品的分离、富集和提纯提供了很大方便 .这些优点使它…     

Chromatographic separation and mass spectrometric identification of positional isomers of polyethylene glycol-modified growth hormone-releasing factor (1-29)

A one-step chromatographic method capable of separating all isomers of polyethylene glycol (PEG)-growth hormone-releasing factor (GRF) (1-29) conjugates was developed. The unmodified GRF (1-29) and seven different isomers of PEG-GRF (1-29) conjugates were separated by using a simple reversed-phase HPLC method depending on the differences of hydrophobicity due to the number and site of PEG attachment. The PEGylation sites of all isomers of PEG-GRF (1-29) conjugates were identified by determining the molecular masses of the Lys-C digested fragments with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. This study is a first report for the separation of all PEG-conjugate isomers and would be useful for further studies to find the promising conjugate by evaluating biological activity and stability of each isomer.     

Dextran-spermine conjugate: an efficient vector for gene delivery

Cationic Polysaccharides based on oligoamine-dextran conjugates were synthesized and tested as vectors for gene transfection. Dextran with 40 kDa in average molecular weight was oxidized under mild conditions by potassium periodate to obtain the respective polyaldehydes in relatively high yields (∼90%). The oxidized dextran was reacted by reductive animation with various oligoamines of 2 to 4 amino groups to obtain the corresponding imine-conjugates. These water-soluble polymers were then reduced by excess of sodium borohydride to obtain the corresponding amine-conjuagtes in 30-40% overall yield. The electrostatic interactions of the representative polycations with plasmid DNA were evaluated as a function of charge ratio (+/−, polymer/DNA) and ionic strength of the medium applying the ethidium-bromide quenching assay. Although most synthetic polycations formed stable complexes with Plasmid DNAs, only the dextran-spermine conjugate of a defined amino content and molecular weight was able to transfect cells with high efficiency.     

Preparation of amylose gel and probe diffusion through the gel membranes

Hydro‐swollen amylose gels were prepared by chemical crosslinking with a polyethylene glycol diglycidyl ether. The degree of swelling of the amylose gel (q_w) increases with increasing the amount of crosslinker in the preparation, which is a result contrary to other cases of chemically crosslinked polymer gels. The diffusion coefficient of a fluorescein and a fluorescein‐dextran conjugate in the gel membranes was determined by the time lag method and increases as q_w of the gels increases.     

Indicator properties of oligoethylene glycol hybrids with sterically hindered phenols

Aqueous solutions of hybrid compounds (conjugates) formed by polyethylene glycols modified at the terminal hydroxy groups with sterically hindered phenol lose their phase stability at a certain temperature which depends on the molecular weight of polyethylene glycol, structure of sterically hindered phenol, conjugate concentration, and composition of the medium. This property may be used to estimate the effect of structural factors on the hydrophobic-hydrophilic balance in the conjugate-water system.     

High-performance size-exclusion chromatographic procedure for the determination of fluoresceinyl isothiocyanate dextrans of various molecular masses in biological media

A high-performance size-exclusion chromatographic procedure, using Nucleosil Diol, for the quantitative analysis of fluoresceinyl isothiocyanate dextrans of various molecular masses (10,000-150,000) in biological media was developed. The influence of the molecular mass and the degree of substitution of the conjugates on the chromatographic behaviour are discussed. In addition to quantitation, the molecular mass of the conjugates with degree of substitution below 1.6 could be estimated from the chromatograms. Linear standard calibration curves were obtained at concentrations down to 0.050 micrograms ml-1 in rabbit plasma and urine and homogenates of rabbit liver, lymph node and muscle when the derivative (degree of substitution 0.85) was monitored by fluorescence detection (lambda ex = 493 nm, lambda em = 520 mm). The fluoresceinyl isothiocyanate dextrans were found to be stable for more than three days at 37 degrees in all biological media under investigation. A pH-rate profile for the alkaline hydrolysis of fluoresceinyl isothiocyanate dextrans was constructed. The applicability of the method to pharmacokinetic studies was demonstrated by recording the plasma concentration-time profile of a fluoresceinyl isothiocyanate dextran T-70 conjugate following intravenous injection to a rabbit. In relation to future pharmacokinetic investigations on dextran conjugates, the results reported indicate that labelling of the parent dextran with fluoresceinyl isothiocyanate and monitoring of the fluoresceinyl isothiocyanate dextran conjugate throughout the organism using the described method is a promising development.     

Improved Pharmacokinetics Properties for Catalase by Site‐Specific Glycosidation with Aminated Dextran

Summary: Catalase was chemically modified with an end‐group aminated dextran derivative via a carbodiimide catalyzed reaction. The enzymatic activity of catalase was increased after glycosidation with 4 mol of polymer. This modification improved the pharmacokinetic behavior of catalase, increasing by 7.8‐ and 20‐fold the plasma half‐life times for the α and β phases, and reducing by 176‐fold the total clearance after intravenous administration in rats.

Schematic of the catalase dextran conjugate synthesized here.     

Glucose Oxidase-dextran Conjugates with Enhanced Stabilities Against Temperature and pH

Multipoint covalent bonding of glucose oxidase (EC 1.1.3.4) to hydrophilic natural polymer dextran and optimization of procedures to obtain, with enhanced temperature and pH stabilities, were studied. Purified enzyme was conjugated with various molecular weight dextrans (17.5, 75, and188 kD) in a ratio of 20:1, 10:1, 1:1, 1:5, 1:10, 1:15, and 1:20. After 1 h of incubation at pH 7, the activities of purified enzyme and conjugates were determined at different temperatures (25°C, 30°C, 35°C, 40°C, 50°C, 60°C, 70°C, and 80°C), and the results were evaluated for thermal resistance. Increases in temperature from 25°C to 50°C did not change the activities of the conjugates. The conjugate, which was prepared with 75 kDa dextran in a molar ratio of 1:5, showed the highest thermal resistance and even the activity still remains at 80°C at pH 7.0. This conjugate also displayed activity in a wide pH range (pH 4.0–7.0) at high temperatures. Conjugate, which was synthesized with 75 kDa dextran in a molar ratio of 1:5, appears to be feasible and useful for biotechnological applications.     

Determination of trimethoprim metabolites including conjugates in urine using high-performance liquid chromatography with combined ultraviolet and electrochemical detection

A high-performance liquid chromatographic method for the determination of trimethoprim metabolites in pig urine was developed. The metabolites-glucuronic acid and sulphuric acid conjugates of phenolic metabolites formed by demethylation of trimethoprim-were quantitated after treatment of urine with beta-glucuronidase (Escherichia coli). The sulphuric acid conjugate was not susceptible to enzymatic hydrolysis and was therefore assayed as the conjugate by use of ion-pair chromatography on the reversed-phase column. In order to find suitable conditions for enzymatic hydrolysis of the glucuronides, the conjugates were obtained by gel chromatography of urine from a [14C] trimethoprim-treated pig.     

Synthesis and Characterization of PEG-scutellarin Conjugates, a Potential PEG Ester Prodrug for the Oral Delivery of Scutellarin

Scutellarin is the active ingredient extracted from Erigeron breviscapus, a traditional Chinese herbal medicine. Pharmacological study showed that scutellarin could significantly reduce the blood viscosity, improve the blood flow, decrease the vascular re…     

Linear oligosulfoxides: Synthesis and solubility studies

Synthesis of three linear oligosulfoxides containing up to six sulfoxide groups was achieved by multiple S_N2 reactions between an alkanethiol and alkyl tosylate to give a linear oligosulfide followed by oxidation of the oligosulfide with sodium periodate to give an oligosulfoxide. The challenge of complete avoidance of partial oxidation and over oxidation was easily overcome using the sodium periodate oxidation conditions. Although sulfoxide is a highly polar functional group, the oligosulfoxides were found to have limited solubility in many solvents including DMSO and water, which disobeys the “like dissolves like” rule. The surprising solubility pattern of oligosulfoxides was discussed in the context of the drastically different solubility patterns of polyethylene glycol (PEG), poly(butylene oxide), and poly(methylene oxide). According to a dissolution model, solubility properties of linear oligomers including the oligosulfoxides and PEGs may be heavily affected by their conformations and the suitability of their conformations in water for maximizing attractive interactions between them and water. Based on these hypotheses, the limited solubility of the present oligosulfoxides may not imply the low solubility of similar molecules.     

Molecular properties of conjugates formed by synthetic hydrophilic polymers and sterically hindered phenols

Water-soluble conjugates are prepared via the chemical modification of poly(vinyl alcohol) and poly(ethylene glycol) by antioxidants taken from the family of sterically hindered phenols. The effects of the degree of substitution of conjugates on the dimensions of molecules and their aggregates are studied by viscometry and light scattering in dilute solutions. It is shown that an increase in the amount of antioxidant groups incorporated into the poly(vinyl alcohol) chain leads to a decrease in the dimensions of single conjugate molecules owing to attraction of hydrophobic groups. Poly(ethylene glycol) molecules carrying end groups of sterically hindered phenols form micellar aggregates that are absent in the solution of the initial polymer. The mean number of molecules involved in such an aggregate is 83. It is found that the presence of hydrophobic end groups in poly(ethylene glycol) molecules causes a sharp reduction in the lower critical solution temperature of a solution relative to that of the initial polymer.     

Functional properties of nisin–carbohydrate conjugates formed by radiation induced Maillard reaction

Nisin–carbohydrate conjugates were prepared by irradiating nisin either with glucose or dextran. Increase in browning and formation of intermediate products was observed with a concomitant decrease in free amino and reducing sugar groups indicating occurrence of the Maillard reaction catalyzed by irradiation. Nisin–carbohydrate conjugates showed a broad spectrum antibacterial activity against Gram negative bacteria (Escherichia coli, Pseudomonas fluorescence) as well as Gram positive bacteria (Staphylococcus aureus, Bacillus cereus). Results of antioxidant assays, including that of DPPH radical-scavenging activity and reducing power, showed that the nisin–dextran conjugates possessed better antioxidant potential than nisin–glucose conjugate. These results suggested that it was possible to enhance the functional properties of nisin by preparing radiation induced conjugates suitable for application in food industry.     

Oriented immobilization of periodateoxidized monoclonal antibodies on amino and hydrazide derivatives of eupergit C

Amino and hydrazyno derivatives of Eupergit C were prepared by reaction of the beads with hexamethylene diamine (HMD) and adipic acid dihydrazide (ADH), respectively. Monoclonal antibodies (mAbs) against carboxypeptidase A (CPA) and horse radish peroxidase (HRP) were prepared, and those that did not inhibit the respective enzymatic activities were selected. The carbohydrate moieties of these antibodies were oxidized by reaction with sodium periodate and then coupled onto the modified beads. The oxidation and coupling reactions were optimized to achieve highly active matrix-conjugated antibodies. Thus, antibody-matrix conjugates that possessed antigen-binding activities close to the theoretical value of 2 mol antigen bound/mol immobilized antibody were obtained.     

Antitumor effect of liposomal histone deacetylase inhibitor-lipid conjugates in vitro

Histone deacetylase inhibitor (HDACI), suberoylanilide hydroxamic acid (SAHA), approved by the Food and Drug Administration (FDA) for the treatment of cutaneous T cell lymphoma, is a promising new treatment strategy for various cancers. In this study, we hypothesized that a liposomal formulation of HDACI might efficiently deliver HDACI into tumors. To incorporate HDACI efficiently into the liposomal membrane, we synthesized six HDACI-lipid conjugates, in which polyethylene glycol(2000) (PEG(2000))-lipid or cholesterol (Chol) was linked with a potent hydroxamic acid, HDACI, SAHA or K-182, by cleavable linkers, such as ester, carbamide and disulfide bonds. Liposomal HDACI-lipid conjugates were prepared with distearoylphosphatidylcholine (DSPC) and HDACI-Chol conjugate or with DSPC, Chol and HDACI-PEG-lipid conjugates, and their cytotoxicities were evaluated for human cervix tumor HeLa and mouse colon tumor Colon 26 cells. Among the liposomes, liposomal oleyl-PEG(2000)-SAHA conjugated with SAHA and oleyl-PEG(2000) via a carbamate linker showed higher cytotoxicity via hyperacetylation of histone H3 and induction of caspase 3/7 activity. These results suggested that liposomal HDACI-lipid conjugates may be a potential tool for cancer therapy.     

Stabilization of horseradish peroxidase by covalent conjugation with dextran aldehyde against temperature and pH changes

Stabilization of Horseradish Peroxidase (HRP; EC 1.11.1.7) against temperature and pH via the formation of the conjugates obtained by multipoint covalent bonding of dextran aldehyde (DA) to the enzyme were studied. Hence, three different molar weighted dextrans (17.5 kD, 75 kD, 188 kD) were covalently bonded to purified enzyme with different molar ratios (n_HRP/n_DA 20/1, 10/1, 1/1, 1/5, 1/10, 1/15, 1/20). The thermal stabilities of the obtained conjugates were evaluated with the activities determined at different temperatures (25, 30, 35, 40, 50, 60, 70, 80°C) applying 60 minutes incubation time. Conjugates formed were characterized by gel-permeation chromatography (GPC) and fluorescence techniques. The conjugate synthesized using dextran 75 kDa with n_HRP/n_DA 1/10 molar ratio showed better thermal stability than other conjugates and purified enzyme at pH 7. This conjugate also has wider activity pH range than purified enzyme. In addition, mentioned conjugate at pH 7 had very long storage lifetime compared to purified enzyme at +4°C and room temperature; which is considered a favorable feature for usage in practice.      

Zinc phthalocyanine labelled polyethylene glycol: preparation, characterization, interaction with bovine serum albumin and near infrared fluorescence imaging in vivo

Zinc phthalocyanine labelled polyethylene glycol was prepared to track and monitor the in vivo fate of polyethylene glycol. The chemical structures were characterized by nuclear magnetic resonance and infrared spectroscopy. Their light stability and fluorescence quantum yield were evaluated by UV-Visible and fluorescence spectroscopy methods. The interaction of zinc phthalocyanine labelled polyethylene glycol with bovine serum albumin was evaluated by fluorescence titration and isothermal titration calorimetry methods. Optical imaging in vivo, organ aggregation as well as distribution of fluorescence experiments for tracking polyethylene glycol were performed with zinc phthalocyanine labelled polyethylene glycol as fluorescent agent. Results show that zinc phthalocyanine labelled polyethylene glycol has good optical stability and high emission ability in the near infrared region. Imaging results demonstrate that zinc phthalocyanine labelled polyethylene glycol can track and monitor the in vivo process by near infrared fluorescence imaging, which implies its potential in biomaterials evaluation in vivo by a real-time noninvasive method.